Human complement receptor 2 (CR2/CD21) is a 145 kiloDalton (“kDa”) transmembrane protein comprised of 15 or 16 short consensus repeat (SCR) extracellular domains, a 28 amino acid single pass transmembrane domain and a short 34 amino acid intracellular domain (1-5). Each of the extracellular SCRs comprises approximately 60-70 amino acid residues and is connected by linker regions of three to eight amino acid residues. All SCRs contain a number of conserved amino acid residues including four cysteine residues, which form a pattern of disulfide bridges connecting Cys1-Cys3 and Cys2-Cys4. CR2 is primarily present on B cells, where it is found in complex with other membrane proteins that promote normal humoral and cellular immune responses (6-9). Using the most distally located (i.e., amino-terminal) SCR domains, SCR1-2, CR2 binds four classes of ligands—complement component 3 (C3) proteolytic fragments iC3b, C3dg and C3d (10, 11); the Epstein-Barr virus (EBV) glycoprotein gp350/220 (gp350) (12-14); the low affinity IgE receptor CD23 (15, 16); and the cytokine interferon alpha (IFNα) (17-19).
The primary role of CR2 is to function as a B cell co-receptor for antigen-mediated B cell activation through enhanced signal transduction (20, 21). This function is carried out through co-ligation via C3d and surface IgM, when C3d is covalently attached to an antigen (22-28). CR2 is also the obligate cellular receptor for EBV through its envelope surface glycoprotein gp350 (12, 20, 29-31). Actual cellular EBV infection is achieved after the ligation of CR2 to gp350 tethers the virus close enough to the cell surface (14, 32, 33), allowing viral gp42 to bind human leukocyte antigen class II molecules (34, 35) and subsequently triggering host cell fusion via three additional viral glycoproteins gB, gH and gL (36-38). IFNα has been shown to be a ligand of CR2, though the physiologic importance of this interaction remains unclear (17-19). It has been suggested, however, that IFNα and CR2 may be involved in the development of the autoimmune disease systemic lupus erythematosus (39-41).
Mutagenesis studies along with structural studies of the CR2-gp350 interaction have suggested residues on CR2 that are required for the interaction (20, 42, 43). ELISA and flow cytometry was used to test candidate CR2 mutants for the binding of gp350 and CR2 (20, 42, 43). In recent studies specific residues on CR2 which were found to have a deleterious effect on gp350-binding when mutated included R13, S15, R28, R36, K41, K57, K67, R83 and R89 (42, 43). In separate work residues P8-S15 within the first conserved inter-cysteine region of SCR1 and the linker region between SCR1 and SCR2 were also highlighted as being essential for gp350-binding to occur (20). These data, in conjunction with separate mutagenesis analyses targeting the gp350 molecule were used to drive an in silico model of the CR2-gp350 interaction utilizing the soft docking program HADDOCK (43-45). This analysis suggested that the primary interaction on CR2 was between SCR1 and the linker region joining SCR1 to SCR2, and for gp350, the linker region between domain 1 and domain 2 (43).
CR2 has been suggested as a receptor for IFNα by the finding that IFNα mimics both gp350 and C3d binding, and the observation that all three ligands bind a similar region on CR2 (18, 19). The mimicry was shown to be functional as well (18). After both the C3d and IFNα structures were solved, the putative CR2 binding sequence was found to have similar structural motifs. IFNα has been described as being able to bind to multiple forms of CR2 from full length to SCR1-2, although to varying degrees (17). Though CR2 has been shown to be a receptor for IFNα, the IFNα binding site within CR2SCR1-2 is unknown.
Further analysis of CR2 interactions with known ligands to identify specific amino acid residues involved in binding to these ligands would enable the design of modified CR2 molecules with defined binding specificity for each known CR2 ligand (e.g., C3 proteolytic fragments iC3b, C3dg and C3d; EBV glycoprotein gp350; CD23; and IFNα.